Frederik Lindberg

Horizontal gene transfer of the Escherichia coli pap and prs pili operons as a mechanism for the development of tissue‐specific adhesive properties

Escherichia coli strains bind to Galα1‐4Gal‐containing glycolipids via P pili‐associated G‐adhesins. Three functional classes of adhesins with different binding specificities are encoded by conserved G‐alleles. We suggest that the Class I papG‐allele of strain J96 is a novel acquisition possibly introduced via horizontal gene transfer into one of the two P pili gene clusters carried by this strain. Closely related strains in the ECOR collection of natural E. coli isolates carry either a Class II or a Class III G‐adhesin. Data indicate that genetic exchanges involving either entire pap or prs gene clusters or individual paplprs genes have occurred. We propose that the retention and spread of pap prs DNA among E. coli is the result of selection pressure exerted by mammalian intestinal isoreceptors.

Uropathogenic, Escherichia coli can express serologically identical pili of different receptor binding specificities

Uropathogenic Escherichia coli frequently express P‐pilus adhesins that recognize Galα(1–4)Gal‐containing glycoconjugates. The P‐pilus adhesin of the, E. coli isolate J96 is encoded by the pap gene cluster and has been shown to agglutinate P1‐erythrocytes. We now describe a novel gene cluster from J96, prs, which is responsible for the agglutination of sheep erythrocytes. The structurally related gene clusters both expressed pili exhibiting the F13 antigen. Analysis of mutants of cloned prs sequences, together with trans‐complementation of pap and prs genes, identified the sheep‐specific adhesin as the 37‐kD PrsG protein. The prsG gene occupies the equivalent position in prs as occupied by papG, which specifies the Galeα(1–4)Gal‐specific adhesion of pap. PrsG was shown to be structurally distinct from PapG since PapG‐specific antiserum did not cross‐react with PrsG. Using a solid phase glycolipid receptor binding assay, PrsG was found to specify preferential binding to the Forssman antigen, a major constituent of sheep erythrocyte membranes. The binding epitope was identified as the GaINAcα(1–3)GaINAc moiety. This is the first direct evidence that serologically identical pili may present antigenically distinct adhesins, each capable of binding to a specific receptor.

Signalling proteins in enterobacterial AmpC β‐lactamase regulation

The cloned Citrobacter freundii ampC β‐lactamase is inducible in the presence of its regulatory gene ampR in Escherichia coli (Lindberg et al., 1985). The basal level of expression and inducibility are affected by two E. coli proteins encoded by the closely linked ampD and ampE genes. Deletion of both genes led to constitutive ampR‐dependent overproduction of β‐lactamase, whereas an out‐of‐frame deletion in AmpD caused the basal expression to increase twofold. This ampD1 mutant was inducible at lower β‐lactam concentrations than the wild type. An IS1 insertion in ampD was polar on ampE expression and increased basal β‐lactamase expression 30‐fold while mediating a semi‐constitutive phenotype. AmpE expressed from a recombinant plasmid in an ampD ampE deletion mutant reduced basal β‐lactamase expression to wild‐type levels but did not markedly reduce β‐lactam resistance since the cells became hyperinducible. in the absence of AmpD, increasing levels of AmpE therefore decrease the basal expression of AmpC β‐lactamase in an AmpR‐dependent manner. AmpD modulated the response exerted on β‐lactamase expression by AmpE. The ampD gene encodes a 20.5kD cytoplasmic protein while the 32.1kD ampE gene product is an integral membrane protein with a likely ATP‐binding site between the second and third putative transmembrane region. Since neither AmpD nor AmpE are needed for β‐lactam induction and since these proteins could not be covalently labelled by benzylpenicillin, they are not thought to act as β‐lactam‐binding sensory tranducers. Instead it is suggested that AmpD and AmpE sense the effect of β‐lactam action on peptidoglycan biosynthesis and relay this signal to AmpR.

Tip proteins of pili associated with pyelonephritis: new candidates for vaccine development

Escherichia coli strains associated with extra-intestinal infections frequently express carbohydrate binding adhesins which are present as minor components of pili. The adhesin protein, PapG, and at least two other minor pilus subunits, PapE and PapF, are associated with the tips of Gal alpha (1-4)Gal-binding Pap pili or P fimbriae of uropathogenic E. coli. The structural and antigenic variation of these tip-associated proteins is discussed and evidence is presented showing that serologically identical pili may contain antigenically distinct adhesins each capable of binding to a specific receptor. One approach to the purification of these tip-associated proteins is presented and involves complex formation with the periplasmic transport protein PapD.

Integrity of Escherichia coli P pili during biogenesis: properties and role of PapJ

The papJ gene of uropathogenic Escherichia coli is required to maintain the integrity of Galα(1‐4)Gal‐binding P pili. Electron microscopy and ELISA have established that strains carrying the papJ1 mutant allele have a large amount of pilus antigen free of the cells. In contrast to the whole pili released by strains unable to produce the PapH pilus anchor, the free papJ1 pili consist of variably sized segments that appear to result from Internal breakages to the pilus. The DNA sequence of papJ is presented and its gene product identified as an 18kD periplasmic protein that possesses homology with nucleotide‐binding proteins. PapJ may function as a ‘molecular chaperone’ directly or indirectly establishing the correct assembly of PapA subunits in the P pilus.

Isolation of the Pre-Assembled Gal α(1–4)Gal-Specific Pilus-Associated Adhesin from the Periplasm in Uropathogenic Escherichia Coli

Escherichia coli is the most common facultative anaerobe found among the normal floral inhabiting the large intestine of healthy individuals. These organisms play a role in maintaining the normal physiological conditions of their environment. However, during the last three decades certain stains of E. coli have come to be recognized as important human pathogens. Enterotoxigenic isolates of E. coli are a primary cause of infant diarrhea and some strains produce haemorrhagic and dysentery-like infections (Levine et al., 1983). In addition to its considerable importance as an intestinal pathogen, E. coli is also recognized as a significant cause of extraintestinal infections (Truck and Petersdorf, 1963). Prevalent nosocomial infections produced include bacteraemia, surgical wound infections and urinary tract infections (Schaeffer and Chmiel, 1983). In the community, E. coli has been primarily associated with urinary tract infections and accounts for up to 80% of all episodes of cystitis and 90% of pyelonephritis cases. E. coli is also responsible for a major proportion of bacterial meningitis in infants (Klein and Marcy, 1976).

Biogenesis of Galα(1–4)Gal binding P-pili of Uropathogenic E. Coli

Escherichia coli is the most common cause of urinary tract infections (UTI). Depending on the virulence of the organism and the efficiency of the host defense system, these UTI appear either as asymptomatic bacteriuria, acute cystitis or acute pyelonephritis. A number of putative virulence properties have been recognized in E. coli isolates associated with acute pyelonephritis in otherwise uncompromised children. One such property is the ability to express mannose-resistant (MR) hemagglutinins. Most E. coli are capable of forming type 1 pili that carry a mannosespecific adhesin. Pyelonephritogenic E. coli additionally carry one or more chromosomal gene clusters that encode pili associated with MR hemagglutination. Of the several MR hemagglutinins that are known today, the best characterized is the Galα(1–4)Gal-specific adhesin associated with P-pili. The name P-pili refers to the observation that this adhesin acts as a hemagglutinin for erythrocytes which express any of the P blood group antigens.